Seven major cyclic nucleotide phosphodiesterase (PDE) gene families with distinct properties have been identified. One class, the Type III (or cGMP-inhibited (cGI)) PDE is characterized by its high affinity for cAMP and cGMP and specific inhibition by certain positive inotropic and vasodilatory agents. cGI PDEs are important in regulation of myocardial contractility, vascular tonus, platelet aggregation and the antilipolytic action of insulin. Our laboratory has cloned cDNAs for two distinct cGI PDE types or subfamilies - cGIP1 (from rat (R) and human (H) adipose tissue) and cGIP2 from rat adipose and human cardiac cDNA libraries. The open reading frames encode proteins of about 122-125 kDa. cGIP1 and cGIP2 PDEs share a common domain organization pattern. The deduced amino acid sequences in the N-terminal portions of the proteins predict regulatory domains, including hydrophobic putative membrane association domains and potential cAMP-dependent protein kinase phosphorylation sites. Collaborative studies indicate that, based on the deduced sequence of RcGIP1, serine 427 in rat adipocyte cGI PDE is a likely target site for phosphorylation by cAMP-dependent protein kinase in vitro. A truncated form of RcGIP1 lacking the N-terminal region hydrophobic domain was expressed in Sf9 insect cells and found to be predominantly cytosolic. The gene for HcGIP1 was localized to Chromosome 11; for HcGIP2, chromosome 12. Expression of RcGIP1 and RcGIP2 mRNAs differs in different tissues. RcGIP1 mRNA is expressed in rat adipocytes from both brown and white adipose tissue and increases during differentiation of 3T3-L1 adipocytes whereas RcGIP2 mRNA is more highly expressed in rat cardiac tissue. Our results indicate that the cGI PDE gene family contains at least 2 subfamilies, products of distinct but related genes. cGIP1 PDEs (or cGIP2 PDEs) from rat and human are more closely related than cGIP1 and cGIP2 PDEs from the same species.